Expression of autophagy-related protein 5 in colon cancer and its impact on the migration and invasion ability of colon cancer cells
Objective To evaluate the expression of autophagy-related protein 5(ATG5)in colon cancer and its rela-tionship with clinicopathological features,and to analyze the effect of ATG5 on the migration and invasion ability of colon cancer cells.Methods The expression levels of ATG5 mRNA and ATG5 protein in colon cancer tissues and adjacent normal tissues were analyzed using the online analysis tool cProsite.Kaplan-Meier Plotter was used to analyze the effect of ATG5 expression level on the prognosis of colon cancer patients.The expression of ATG5 in 100 colon cancer tissues was detected by immunohistochemistry,and the correlation between the expression level of ATG5 and clinicopathological features was analyzed by χ2 and Fisher's test.ATG5 was overexpressed or knocked down in colon cancer cells by transfecting plasmid or small interfering RNA(siRNA),and then the transfection efficiency and changes of microtubule-associated protein light chain 3(LC3)were detected by Western blotting.Transwell assay was used to evaluate migration and invasion ability of colon cancer cells after overexpression or knockdown.Differentially expressed genes associated with ATG5 in colon cancer were analyzed using the LinkedOmics database,and gene ontology(GO)and kyoto encyclopedia of genes and genomes(KEGG)enrichment analyses were performed.Results The expression levels of ATG5 mRNA and protein in colon cancer tissues were lower than those in adjacent normal tissues(P<0.001).The recurrence-free survival of colon cancer patients with low ATG5 expression was significantly shorter than those with high ATG5 expression(P<0.001).Immunohistochemistry showed that the expression level of ATG5 in colon cancer tissues was correlated with lymph node metastasis,and low expression of ATG5 in colon cancer patients was more likely to develop lymph node metastasis(P=0.027).Western blotting and Transwell experiments showed that after overex-pression of ATG5 in SW1116 cells,the expression of LC3Ⅱ/LC3Ⅰ increased,and the migration and invasion ability of cells was weakened(PATG5=0.001;PLC3Ⅱ/LC3Ⅰ=0.04;Pmigration<0.001,Pinvasion<0.001).Conversely,ATG5 knock-down in DLD1 cells led to a decrease in the expression of LC3Ⅱ/LC3Ⅰ,and an increase in the migration and invasion ability of the cells(PATG5#1=0.021,PATG5#2<0.001;PLC3Ⅱ/LC3Ⅰ#1=0.013,PLC3Ⅱ/LC3Ⅰ#2=0.02;Pmigration<0.001,Pinvasion<0.001).ATG5-related differentially expressed gene enrichment analysis suggested that ATG5 might affect colon cancer metastasis by affecting DNA damage response,chromatin organization and Notch signaling pathway.Conclusion ATG5 is lowly expressed in colon cancer and is significantly correlated with lymph node metastasis and prognosis of patients.ATG5 in colon cancer cells can increase the level of autophagy and inhibit cell migration and invasion,sugges-ting that ATG5 and its regulated autophagy process may become a new target for the clinical treatment of colon cancer.
Autophagy-related protein 5Tumor metastasisAutophagyColon cancerMicrotubule-associated protein light chain 3
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